title: Gamma subunit second transmembrane domain contributes to epithelial sodium channel gating and amiloride block.
abstract: The epithelial sodium channel (ENaC) is comprised of three homologous subunits. Channels composed solely of α- and β-subunits (αβ-channels) exhibit a very high open probability (Po) and reduced sensitivity to amiloride, in contrast to channels composed of α- and γ-subunits or of all three subunits (i.e., αγ- and αβγ-channels). A mutant channel comprised of α- and β-subunits, and a chimeric γ-subunit where the region immediately preceding (β12 and wrist) and encompassing the second transmembrane domain (TM2) was replaced with the corresponding region of the β-subunit (γ-βTM2), displayed characteristics reminiscent of αβ-channels, including a reduced amiloride potency of block and a loss of Na(+) self-inhibition (reflecting an increased Po). Substitutions at key pore-lining residues of the γ-βTM2 chimera enhanced the Na(+) self-inhibition response, whereas key γ-subunit substitutions reduced the response. Furthermore, multiple sites within the TM2 domain of the γ-subunit were required to confer high amiloride potency. In summary, we have identified novel pore-lining residues of the γ-subunit of ENaC that are important for proper channel gating and its interaction with amiloride.
authors.affiliation: Renal-Electrolyte Division, A919 Scaife Hall, 3550 Terrace St., Pittsburgh, PA 15261. kleyman@pitt.edu.
authors.forename: Shujie
authors.initials: S
authors.lastname: Shi
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Kleyman
mesh_terms: D000584:Amiloride / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000818:Animals; D053503:Epithelial Sodium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015640:Ion Channel Gating; D017136:Ion Transport / Q000187:drug effects* / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D018408:Patch-Clamp Techniques; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014981:Xenopus
chemical_list: D053503:Epithelial Sodium Channels; D021122:Protein Subunits; D000584:Amiloride
keywords: ENaC; amiloride; open probability; sodium; subunit